Quantitative analysis of amitriptyline and nortriptyline in human plasma and liver microsomal preparations by high-performance liquid chromatography

A simple, rapid, highly selective and sensitive method for the analysis of amitriptyline and nortriptyline in plasma and human liver microsomes is described. It is suitable for the routine analysis of large numbers of samples using readily available instrumentation and low cost consumables. The detection limit was 2 ng/ml for both compounds and calibration curves were linear over a wide range of concentrations and passed through the origin. The within-batch and between-batch coefficients of variation for amitriptyline and nortriptyline were less than 7.4% and 12.8%, respectively. A series of compounds, including inhibitors used for probing cytochrome P450 activity in vitro, were tested for interference in the assay. Only ketoconazole caused interference and the assay was modified to allow samples containing ketoconazole to be analysed.     

Sensitive and rapid method for the simultaneous quantification of five antidepressants with their respective metabolites in plasma using high-performance liquid chromatography with diode-array detection

A high-performance liquid chromatographic method with diode array detection (HPLC-DAD) has been developed for the simultaneous separation and quantification of imipramine, amitriptyline, maprotiline, fluoxetine, clomipramine and their respective metabolites using a 500-μl plasma sample and clovoxamine as the internal standard. The substances were eluted on a Symmetry C₁₈ 5-μm column (250×4.6 mm, I.D.). Full UV spectra from 200 to 450 nm were recorded on-line during the entire analysis and were automatically compared to spectra stored in a library. The quantification was performed at 226, 254, and 400 nm. Peak height ratios were linear over a concentration range of 10–3000 ng/ml: the correlation coefficient (r) was better than 0.998 for all substances at each wavelength. Acceptable coefficients of variation are demonstrated for both within-run and day-to-day assays. The method is simple, highly specific and currently being used for drug monitoring and toxicological studies in children and adult patients.